CDCA3在肝细胞肝癌增殖中的作用研究

Study on the role of CDCA3 in the proliferation of hepatocellular carcinoma

目的探讨细胞周期相关因子3(CDCA3)在肝癌中的生物学行为。方法收集2018年1月~2019年6月在武汉大学人民医院肝胆外科行手术治疗的肝细胞肝癌患者的组织标本,共30例肝癌标本和癌旁标本;分别采用Real-Time PCR检测不同组织中的CDCA3的表达情况;采用lip3000转染法分别将CDCA3干扰RNA序列和对照序列转入肝癌细胞株,应用CCK-8、EdU免疫荧光染色检测细胞增殖,流逝细胞术检测细胞周期。结果肝癌组织CDCA3的表达量明显高于癌旁组织,差异有高度统计学意义(P<0.01)。HepG2、Huh7、SMMC-7721细胞中CDCA3的表达量明显高于LO2细胞,差异有统计学意义(P<0.05或P<0.01),而BEL-7402细胞中CDCA3的表达量与LO2细胞的表达量比较,差异无统计学意义(P>0.05)。siCDCA3#1和siCDCA3#2下调SMMC-7721细胞CDCA3的表达量与对照组(siNC)比较,差异均有统计学意义(均P<0.05),故选取SMMC-7721细胞完成后续研究。siCDCA3#1和siCDCA3#2下调SMMC-7721细胞CDCA3的表达量后,24 h和48 h细胞增殖数明显低于siNC,差异均有统计学意义(均P<0.05)。并且EdU检测阳性细胞数明显低于siNC。siCDCA3#2下调SMMC-7721细胞CDCA3表达量后,细胞周期的G1期时间较siNC延长,差异有统计学意义(P<0.05)。结论CDCA3能够促进肝癌细胞增殖,是潜在的治疗靶点。

Objective To investigate the biological behavior of cell division cycle associated protein 3(CDCA3)in liver cancer.Methods Tissue specimens of hepatocellular carcinoma patients who underwent surgical treatment in the Department of Hepatobiliary Surgery,Renmin Hospital of Wuhan University from January 2018 to June 2019 were collected.There were 30 liver cancer specimens and paracancer specimens.Real-time PCR was used to detect the expression of CDCA3 in different tissues.CDCA3 interfering RNA sequences and control sequences were transfected into hepatocellular carcinoma cell lines by lip3000 transfection.Cell proliferation was detected by CCK-8 and EdU immunofluorescence staining,and cell cycle was detected by elapse cytology.Results The expression level of CDCA3 in liver cancer tissues was significantly higher than that in paracancer tissues,and the difference was highly statistically significant(P<0.01).The expression levels of CDCA3 in HepG2,Huh7 and SMMC-7721 cells were significantly higher than those in LO2 cells,with statistically significant differences(P<0.05 or P<0.01),while the expression level of CDCA3 in BEL-7402 cells was not statistically significant compared with that in LO2 cells(P>0.05).The expression of CDCA3 in SMMC-7721 cells down-regulated by SiCDCA3#1 and siCDCA3#2 was statistically significant compared with that in the control group(siNC)(P<0.05).Therefore,SMMC-7721 cells were selected to complete the follow-up study.After the expression levels of CDCA3 in SMMC-7721 cells were down-regulated by siCDCA3#1 and siCDCA3#2,the number of cell proliferation at 24 h and 48 h was significantly lower than that in the siNC,and the differences were statistically significant(all P<0.05).And the number of positive cells detected at EdU was significantly lower than that in the siNC.Compared with the siNC,the G1 phase time of SMMC-7721 cell cycle was prolonged after the expression of CDCA3 in SMMC-7721 cell was down-regulated by siCDCA3#2,and the difference was statistically significant(P<0.05).Conclusion CDCA3 can promote the proliferation of liver cancer cells and is a potential therapeutic target.